At the present time many medicaments are in development which are intended to combat tumors and their metastases which spread in bone. Despite all advances in medicaments, bone metastases, however, do not count as being curable or treatable. There are attempts, by means of antibodies against surface antigens of tumor cells, to combat their metastases. Bone metastases are, however, despite this the cause of death in 73% of cases of tumors of the breast, and 68% of cases for tumors of the prostate gland. For tumors of other tissues the following figures apply: cervix 50%, thyroid gland 42%, bladder 40%, lungs 36%, ovaries 9% and colon 6%.
Tumor cells which express the so-called bone sialoprotein have the peculiarity that they prefer to settle in bony tissue and form metastases there, particularly in the case of tumors of the prostate gland, breast, lungs, kidney and thyroid and less frequently in the case of malign and semi-malign tumors. The bone sialoprotein (BSP) is a phosphorylated bone glycoprotein having a relative mass of ca. 80 kDa in the SDS-PAGE. The cDNA for BSP codes for a peptide sequence of ca. 33 kDa (Fisher L. W. et al. (1990), J. Biol. Chem. 265, 2347-51; U.S. Pat. No. 5,340,934). BSP is one the few matrix proteins the occurrence of which on mineralising tissue such as bones, dentin and calcifying cartilage is restricted. BSP represents ca. 10 to 15% if the total non-collagenic proteins in the bone matrix. It is as a rule expressed by cells which take part in the formation of dentin, bones and cartilage, for example osteoblasts, developing osteocytes, hypertrophic chondrocytes, odontoblasts and cementoblasts, but also by the trophoblasts in the placenta and some types of cancer cells, e.g. in the case of lungs, breast, prostate, kidney, thyroid and neuroblastoma primary and secondary tumors, in the case of multiple myeloma and in bone metastases. The degree of expression of BSP by the tumor closely correlates with the severity of the cancer (Waltregny D. et al., Increased expression of bone sialoprotein in bone metastases compared with visceral metastases in human breast and prostate cancers, in J. Bone Miner. Res., 2000, 15(5), 834-43; Bellahcéne, A. et al., Bone sialoprotein expression in primary human breast cancer is associated with bone metastases development, in J. Bone Miner. Res., 1996, 11, 665-670; Waltregny, D. et al., Prognostic value of bone sialoprotein expression in clinically localised human prostate cancer, in Journal of the National Cancer Institute, 1998, 90, 1000-1008; Bellahcéne, A. et al., Expression of bone sialoprotein in primary breast cancer is associated with poor survival, in Int, J. Cancer, 1996, 69, 350-353).
BSP, as an adhesion molecule, is supposed to bring about attachment and dissemination of cells on the tissue matrix, since in vitro it forms crystallisation nuclei for biological apatite and in vivo takes part in mineralisation. The switching off of the BSP gene in knock-out mice leads to no recognisable disruption of the building and functioning of the skeleton. In tumors BSP is attributed with participation in microcalcification (Castronovo, V. et al., Evidence that breast cancer associated microcalcifications are mineralized malignant cells, in Int. J. Oncol., 1998, 12, 305-308) and the colonisation of bones by metastasising tumor cells (Bellahcène, A. et al., Expression of bone sioloprotein in primary breast cancer is associated with poor survival, in Int. J. Cancer, 1996, 69, 350-353).
The level of concentration of BSP in the serum of patients with primary carcinomas serves for diagnosis of whether these patients have bone metastases or such are likely to arise from the primary tumor (Diploma Thesis of Ms. Ina-Alexandra Meier, Development of a radioimmunoassay for the determination of bone sialoprotein (BSP) [“Entwicklung eines Radioimmunoassays zur Bestimmung von Bonesialoprotein (BSP)]”, 1996, Darmstadt, Technical University [Fachhochschule], Specialist Field Chemical Technology [FB Chemische Technologie]; Dissertation of Mr. Markus Karmatschek, Isolation of bone sialoprotein from human bones, Structure of a radioimmunoassay for the measurement thereof in serum [“Isolierung von Bonesialoprotein aus humanem Knochen, Aufbau eines Radioimmunoassays zur dessen Messung im Serum”], 1996; Specialist Field of Biology of the Technical University of Darmstadt [FB Biologie der Technischen Hochschule Darmstadt]; Diel I. J. et al., Elevated bone sialoprotein in primary breast cancer patients is a potent marker for bone metastases; in Proceedings of ASCO, 1998, 17, Abstract 461; Diel I. J. et al, Serum bone sialoprotein in patients with primary breast cancer is a prognostic marker for subsequent bone metastasis, in Clin. Cancer Res., 1999, 5, 3914-19; DE 198 13 633; DE 198 21 533; WO 99/50666).
However, in body fluids free BSP is bound by complement factor H with high affinity. Further, BSP can bind to various receptors. Thus, there have been produced in rabbits antibodies against various peptide partial structures of BSP (Fisher, L. W. et al., Antisera and cDNA probes to human and certain animal model bone matrix noncollagenous proteins. Acta Orthop Scand Suppl., 1995, 266, 61-655), against recombinant BSP (Stubbs J T 3rd et al., Characterization of native and recombinant bone sialoprotein: delineation of the mineral-binding and cell adhesion domains and structural analysis of the RGD domain. J. Bone Miner. Res. 1997 12(8), 1210-22), and against BSP isolated from bones, which antibodies failed to recognise any BSP in human serum. The larger factor H molecule of 150 kDa probably masks the smaller BSP (of ca. 65 kDa), so that antibodies cannot bind. Further, factor H is present in excess in the serum (0.5 mg factor H/mL in comparison to BSP with <20 ng/ml Serum in the case of healthy persons and max. 160 ng/ml in the case of tumor patients). It has been asserted that immunological direct determination of BSP in body fluids is impossible, without reducing sample preparation, due to the binding to the factor H and possibly that trophoblasts and BSP producing tumor cells are thereby protected from attack by the immune system, since the factor H belongs to the complement system and is known to bring about a restriction of the alternative pathway to complement lysis (Fedarko N. S. et al., Factor H binding of bone sialoprotein and osteopontin enables tumor cell evasion of complement-mediated attack, in J. Biol. Chem., 200, 275, 16666-16672; WO 00/062065). Further, BSP can specifically bind to the integrin receptors on the cell surface through its own recognition sequence (arginine-glycine-aspartate, RGD). In the case of expression of BSP the tumor cells are then supposed to bind the factor H in the blood and in the tissue fluids to their cell surfaces, or concentrate it around them. Such a protection of BSP from the complement system of the blood of the mother is suspected also for the trophoblasts in the placenta (Fedarko N. S. et al. Factor H binding of bone sialoprotein and osteopontin enables tumor cells evasion of complement-mediated attack, in J. Biol. Chem., 200, 275, 16666-16672; WO 00/062065). Further there is also suspected a function of BSP in angiogenesis. Along with the adhesion of osteoclasts and osteoblasts to the bone matrix—through the binding of the RGD recognition sequence in the matrix to the alpha(v)beta(3) integrin receptors on the cell wall—it is also observed that the adhesion, dissemination and orientation of the endothelial cells is probably mediated by BSP. Namely, blood vessel formation around a tumor occurs in parallel with the BSP expression in the tumor cells (Bellahcène A et al., Bone sialoprotein mediates human endothelial cell attachment and migration and promotes angiogenesis, in Circ. Res. 2000, 86(8), 885-91).
These characteristics thus make BSP a starting point for medicaments of all kinds. Thus, the binding of BSP via the RGD sequence to vitronectin or integrin receptors of tumor and epithelial cells can be restricted by antagonists (U.S. Pat. No. 6,069,158; U.S. Pat. No. 6,008,213; U.S. Pat. No. 5,849,865; van der Pluijm et al., Bone Sialoprotein peptides are potent inhibitors of breast cancer cell adhesion to bone in vitro, in Cancer Res., 1996, 56, 1948-1955). EP 1 084 719 A1 teaches a pharmaceutical composition having BSP as active substance for the support of the repair of damaged bone and connective tissue. WO 94/13310 teaches a composition having a BSP binding protein of Staphylococcus aureaus as active ingredient. WO 00/36919 discloses regulatory elements for the purposive monitoring and suppression of the expression of BSP in tumor and connective tissue cells, which promote calcification. Thus, generally the substances of the regulation of the cell growth and cell migration are of particular interest from a diagnostic and therapeutic point of view. There are, however, still very many unknown factors which control cancer growth, whereby primary and secondary tumors and colonised organs interact. Here, important steps are the invasion, adhesion, migration and cell division of the tumor cells. Along with matrix metalloproteinases, adhesion molecules and chemotactic factors play a particular role. A medicament for combating and also for healing bone metastases on the basis of antibodies and binding molecules against BSP is not known. It is also not known that the BSP of tumor cells is different from the BSP of normal healthy cells.